Bacterial regulation of intestinal antimicrobial defense

描述（由申请人提供）：哺乳动物宿主面临的一个主要挑战是与定植在肠道中的大量细菌群落保持共生关系。肠上皮是肠道微生物群与宿主内部组织之间的主要屏障，但上皮细胞如何控制细菌与粘膜表面的相互作用并限制微生物对肠道屏障的渗透，目前尚不清楚。我们发现了一种新的抑菌因子RegIIIg，它是由小肠上皮细胞产生的。RegIIIg是碳水化合物结合蛋白c型凝集素家族的一员，可排入肠腔。RegIIIg对革兰氏阳性菌具有直接杀菌活性，并通过与肽聚糖碳水化合物的相互作用结合其细菌靶标。因此，RegIIIg代表了一种以前未被认识的粘膜防御机制和c型凝集素的一种新的生物学功能。肠道细菌通过直接激活上皮细胞toll样受体来调节RegIIIg转录，揭示了肠道细菌和肠上皮之间调节抗菌活性的直接对话。由于其他几种Reg家族c型凝集素是由肠道上皮产生的，因此直接杀菌的c型凝集素可能代表了肠道上皮防御的重要一般机制。本建议的总体目标是扩展我们对凝集素介导的肠粘膜表面抗菌防御的理解。这项工作将涉及三个具体目标。首先，我们将使用体外生化方法来确定凝集素介导的杀菌活性的分子机制。其次，我们将使用RegIIIg敲除小鼠来描述RegIIIg如何在体内促进肠道宿主-微生物稳态。第三，我们将在体外和体内分析其他Reg蛋白，以探索直接杀菌凝集素是否构成粘膜防御的一般机制。总之，这些研究将导致对细菌-粘膜关联的免疫控制的基本见解，并将为共生宿主-微生物关联提供新的视角。

DESCRIPTION (provided by applicant): A major challenge for mammalian hosts is to maintain symbiotic relationships with the vast bacterial communities that colonize the intestines. The intestinal epithelium is the primary barrier between the intestinal microbiota and internal host tissues, yet little is known about how epithelial cells control bacterial interactions with the mucosal surface and limit microbial penetration of the intestinal barrier. We have discovered a novel antibacterial factor, RegIIIg, which is produced by small intestinal epithelial cells. RegIIIg is a member of the C-type lectin family of carbohydrate-binding proteins that is discharged into the gut lumen. RegIIIg has direct bactericidal activity against Gram-positive bacteria and binds its bacterial targets via interactions with peptidoglycan carbohydrate. RegIIIg thus represents a previously unappreciated mechanism of mucosal defense and a new biological function for C-type lectins. Intestinal bacteria regulate RegIIIg transcription through direct activation of epithelial cell Toll-like receptors, revealing a direct dialog between enteric bacteria and the intestinal epithelium that regulates antimicrobial activity. As several other Reg family C-type lectins are produced by gut epithelia, directly bactericidal C-type lectins likely represent an important general mechanism of gut epithelial defense. The overall goal of this proposal is to extend our understanding of lectin- mediated antimicrobial defense of the intestinal mucosal surface. This work will involve three Specific Aims. First, we will use in vitro biochemical approaches to determine the molecular mechanism of lectin-mediated bactericidal activity. Second, we will use RegIIIg knockout mice to delineate how RegIIIg contributes to intestinal host-microbial homeostasis in vivo. Third, we will analyze other Reg proteins in vitro and in vivo in order to explore whether directly bactericidal lectins constitute a general mechanism of mucosal defense. Together, these studies should lead to fundamental insights into immune control of bacterial-mucosal associations, and will provide new perspectives on symbiotic host-microbial associations. PUBLIC HEALTH RELEVANCE: A major challenge for humans is to maintain symbiotic relationships with the vast bacterial communities that colonize the intestines. Intestinal epithelial cells produce abundant quantitites of RegIIIg, a carbohydrate binding protein that has a remarkable capacity to kill bacteria. In this grant we propose to explore the mechanism that RegIIIg uses to kill bacteria, and to understand how this protein functions to protect deeper host tissues from invasion by the intestine's normal microbial inhabitants. The results from these studies should yield new strategies for designing novel antimicrobial therapeutics, and could lead to new approaches to treating inflammatory bowel disease.